There are eight known Transmembrane channel-like (TMC) isoforms in the human TMC gene family (TMC1-8), several of which are linked to inherited human diseases. The disease linkages, and the contrasting and generally wide tissue distribution of the TMC genes, indicate the importance of this gene family to human physiology and pathophysiology. Yet, despite numerous studies, the precise function of mammalian TMC proteins, which share no homology with other gene families, has remained enigmatic. Recent elegant studies relying upon recordings in auditory hair cells from mice with Tmc1 and/or Tmc2 genes deleted strongly suggest a role in hair cell mechanotransduction, explaining the linkage of these genes to inherited deafness. However, despite great interest, direct functional analysis of mammalian TMC gene function has been hampered by an inability to express TMC proteins at the cell surface in heterologous expression systems, to facilitate study of their putative roles as ion channels or their regulatory subunits. The TMC gene family therefore constitutes a rare entity: an orphan gene family of uncertain function, with several established disease linkages. Seeking to address this major gap in knowledge, we hypothesized that TMC proteins require other proteins to reach the cell surface. We developed a simple surface expression screen and discovered that TMC1 surface expression is specifically rescued by the KCNQ1 voltage-gated potassium (Kv) channel ? subunit. We have also discovered that TMC1 inhibits the typical KCNQ1 current, instead forming a new current with novel attributes. The data define human TMC1 as a novel type of channel subunit, paving the way for long-awaited functional studies of mammalian TMC genes. We now propose to define mechanisms of TMC1 function and pathobiology, and to open up the rest of the TMC family to functional study by us and other groups, as quickly as possible, to facilitate future discovery of therapeutic approaches for TMC-linked human diseases. In Aim 1, we will test fundamental hypotheses regarding the functional role of TMC1 in complexes with KCNQ1, determining how TMC1 is activated and which functional properties of KCNQ1 are altered by TMC1 and vice versa. In Aim 2, with an innovative application of a high-throughput surface exposure assay followed by two different high-throughput functional assays (fluorescence-based and electrophysiological) we will test the hypothesis that all eight TMC proteins are ion channel subunits, and that they form complexes with other members of the forty-strong Kv ? subunit gene family. The overall goal is to discover the basic functional mechanistic attributes of one of the few remaining enigmatic disease-linked human ion channel gene families.